Effects of melatonin on behavioral dopaminergic supersensitivity

This study examines the effects of melatonin on dopaminergic supersensitivity induced by long-term treatment with haloperidol in rats. Enhancements of spontaneous general activity in an open-field and of stereotyped behavior induced by apomorphine after abrupt withdrawal from long-term treatment with haloperidol were used as experimental parameters for dopaminergic supersensitivity. Experiment 1 was conducted to investigate the effects of melatonin on the development of dopaminergic supersensitivity, and experiment 2 was conducted to investigate the effects of melatonin on the development as well as on expression of dopaminergic supersensitivity. Rats of both experiments were long-term treated with saline or haloperidol concomitant to saline or melatonin. In experiment 1 behavioral observations were performed after abrupt withdrawal from long-term treatment. In experiment 2 behavioral observations were performed 1 hour after an acute injection of saline or melatonin, administered after the abrupt withdrawal from long-term treatment. Both behavioral parameters used showed the development of central dopaminergic supersensitivity in rats treated with haloperidol since 24 hours after abrupt withdrawal. Concomitant treatment with melatonin intensified haloperidol-induced dopaminergic supersensitivity, observed 72 hours after withdrawal. Melatonin treatment per se also induced behavioral supersensitivity evaluated by both open-field and stereotyped behaviors, although it was more fugacious than that presented by haloperidol. Acute treatment with melatonin reverted the enhancement of the haloperidol-induced dopaminergic supersensitivity produced by concomitant long-term treatment with melatonin, as well as melatonin-induced dopaminergic supersensitivity per se. Our results support previous evidence of antidopaminergic effects of melatonin and demonstrate that repeated administration of this hormone modifies the plasticity of behaviors mediated by central dopaminergic systems.     

Perlecan-enriched intercellular space of junctional epithelium provides primary infrastructure for leukocyte migration through squamous epithelial cells

The aim of this study was to demonstrate the presence of intraepithelial stroma represented by extracellular matrix (ECM) deposits in the junctional epithelium to clarify its function as a scaffold for leukocyte migration through epithelial cells. Twenty-three biopsy specimens from the gingiva including the junctional epithelium were examined to determine comparative protein and gene level expression profiles for keratin and ECM molecules between the junctional epithelium and the gingival epithelium using immunohistochemistry and in situ hybridization. Intraepithelial leukocyte types and frequencies were also determined and compared between the junctional and gingival epithelia. In the junctional epithelium, which was positive for keratin 19, perlecan was strongly deposited in intercellular space of the whole epithelial layer, while it was faintly positive around the parabasal layer of the gingival epithelium. Perlecan mRNA signals were enhanced to a greater degree in both epithelial and inflammatory cells within the junctional epithelium. In the junctional epithelium, greater numbers of neutrophils and macrophages were found as compared with the gingival epithelium. Our results showed that perlecan is the primary ECM molecule comprising intraepithelial stroma of the junctional epithelium, in which leukocytes may migrate on ECM scaffolds in intercellular space toward the surface of the gingival sulci or pockets.     

Hydrology of Dutch Cirsio‐Molinietum meadows: Prospects for restoration

Abstract. Fen meadows (Cirsio dissecti‐Molinietum) are seriously threatened by desiccation, acidification and eutro‐phication. In The Netherlands several projects were launched to restore damaged fen meadows. This review describes how successes and failures of these restoration projects depend on hydrological systems. Six hydrological systems have been distinguished, which all provide the site conditions required by this community. Nowadays, the best developed fen meadows are found in the higher Pleistocene landscape of The Netherlands, where they depend on base‐rich groundwater discharging from local or large groundwater systems. Fen meadows of the lower Holocene landscape usually occur in man‐made surface water systems. Almost all stands have been severely deteriorated. Restoration of fen meadows in the Pleistocene landscape is promising when the hydrology is only slightly disturbed or when hydrological measures are taken in combination with sod cutting. Restoration prospects of fen meadows in the Holocene landscape are low. Until now a complete regeneration of Cirsio‐Molinietum meadows has not been realized. Restoration measures failed to restore high pH values in the top soil. It is hypothesized that viable seeds of many target species lack in the soil seed bank. In addition, the dispersal capacities of these species seem to be limited.     

The effects of drainage on groundwater quality and plant species distribution in stream valley meadows

Conditions in fen meadows in Dutch stream valleys are influenced by both deep (Ca²⁺-rich) and shallow (Ca²⁺-poor) groundwater flows. The distribution patterns of phreatophytic (groundwater-influenced) plant species showed distinct relationships with the distribution of different groundwater types. Large fluctuations in the ionic composition of groundwater were observed in the upper peat layers of drained areas. Ca²⁺-rich groundwater was replaced by precipitation water to a considerable depth (1–1.5 m). These fluctuations in groundwater composition were less pronounced in undrained areas.It was observed that characteristic species of fen meadow communities and rare phreatophytic species were restricted to areas with high water tables, which were nourished by either Ca²⁺-poor or by Ca²⁺-rich groundwater. Few species showed a preference for drained areas, where replacement of groundwater types gave rise to the occurrence of an intermediate groundwater type, which was thought to be the result of an increased acidification of the top soil (increased influence of infiltration water).It was argued that the endangered species were best preserved in areas with an undisturbed discharge of natural groundwater flows.Abbreviations EC = Electrical conductivity     

Isolation of cDNA for a Xenopus sperm-specific basic nuclear protein (SP4) and evidence for expression of SP4 mRNA in primary spermatocytes

A cDNA library was prepared in lambda gt 11 from poly(A)+ mRNA isolated from a pure population of Xenopus round spermatids and screened with an antibody against SP3-5 (sperm-specific proteins) of Xenopus sperm. Positive clones were sequenced and an arginine-rich clone, designated pXSP531, was obtained. The 473-nucleotide sequence of pXSP531 contained an open reading frame of 237 nucleotides which was preceded by a 5' untranslated region of 67 nucleotides. The 3' untranslated region contained 149 nucleotides, including a consensus polyadenylation signal (AAATAAAA). Twenty nucleotides of a poly(A) tail was contained in the pXSP531. SP3-5 were separated from each other by reverse-phase chromatography and sequenced. The amino acid sequence of the peptide fragments which were obtained by digestion of SP4 with V8 protease and separated by reverse-phase chromatography was identical to the sequence of the N-terminal 43 and C-terminal 15 amino acids deduced from the nucleotide sequence of pXSP531. This result demonstrates that pXSP531 encodes SP4. Northern hybridization of RNA extracted from primary spermatocytes and round spermatids on Days 0 and 6 with SP4 cDNA probe (pXSP531) showed that SP4 mRNA is present both in primary spermatocytes and in round spermatids as is protamine mRNA in the rainbow trout. The size of the SP4 mRNA in round spermatids on Day 0 was longer by 60 nucleotides compared to that in primary spermatocytes and that in spermatids on Day 6 was shorter by 30 nucleotides compared to that on Day 0. These size differences were due to differences in the length of the poly(A) tracts because digestion of poly(A) with ribonuclease H resulted in the shortening of mRNA to the same size for three stages.     

Synthesis of sperm-specific basic nuclear proteins (SPs) in cultured spermatids from Xenopus laevis

The accumulation and synthesis of sperm-specific basic nuclear proteins (SPs) in Xenopus spermatids in vitro were studied by acid-urea-Triton polyacrylamide gel electrophoresis and fluorography. In synchronous cultures of round spermatids, the amount of SP2 and SP3-5 accumulated almost linearly with time, while that of SP1 remained almost constant. Fluorography showed that round spermatids incorporated [14C]arginine mostly into SP1 and SP3-5, very little into SP2, and none into histones. When [14C]arginine was incorporated into cells for 24 h on Days 0, 3, and 6, followed by immediate extraction of basic nuclear proteins, the SP1 band was detected faintly on Day 0 and the intensity increased to the maximum level by Day 3 and remained constant on Day 6; the SP3-5 bands were first detected on Day 3 and their intensity increased by Day 6. Thus, SP1 and SP3-5 were synthesized differentially during the culture period. When [14C]arginine or [14C]lysine was incorporated into round spermatids on Days 0, 3, and 6 for 15 h and chased for 3-12 days, the intensity of the SP2 band increased significantly, while the intensity of the SP1 band decreased concomitantly. This result indicates that SP2 was processed from a precursor protein which is probably SP1.     

Molecular mapping around the centromere of tomato chromosome 6 using irradiation-induced deletions

 Irradiation-induced deletion mapping was exploited to construct a detailed locus-order map around the centromere of tomato chromosome 6 (CEN  6). An F₁ hybrid heterozygous for the marker loci thiamineless (tl), yellow virescent (yv) and potato leaf (c), and homozygous recessive for the nematode resistance gene mi, was pollinated with γ-irradiated pollen from cultivar VFNT Cherry carrying the wild-type alleles at the corresponding loci. A dose of 100 Gy was found optimal for inducing mutants. By screening for pseudo-dominant plants showing the marker phenotypes and/or nematode susceptibility, 30 deletions encompassing one or more of the four loci were detected in the M₁ generation. Molecular-marker analysis revealed that 29 of these mutants included the tl and mi loci on the short arm and originated from terminal deletions of different sizes. Remarkably, the breakpoints of these deletions were not randomly distributed along the short arm but located within the centromeric heterochromatin. Only one yv interstitial deletion and no c mutations on the long arm of the chromosome were detected. Mapping of the various chromosomal breakpoints in the isolated mutants permitted the resolution of a cluster of molecular markers from the centromeric heterochromatin that was hitherto unresolvable by genetic linkage analysis. The usefulness of such a deletion-mapping approach for whole-genome mapping is discussed. Received: 4 March 1997 / Accepted: 2 June 1997     

Nutrient limitations in an extant and drained poor fen: implications for restoration

In a species-rich poor fen (Caricetum nigrae) and a species-poor drained fen, the difference in nutrient limitation of the vegetation was assessed in a full-factorial fertilization experiment with N, P and K. The results were compared to the nutrient ratios of plant material and to chemical analysis of the topsoil. A rewetting experiment with intact sods was carried out in the glasshouse and the results are discussed in view of restoration prospects of drained and degraded peatlands. In the undrained poor fen the above-ground biomass yield was N-limited while the vegetation of the drained fen was K-limited. Experimental rewetting of intact turf samples, taken in the drained site, did not change the biomass yield or the type of nutrient limitation. It was concluded that mire systems which have been subjected to prolonged drainage are inclined to pronounced K-deficiency, probably due to washing out of potassium and harvesting the standing crop. This may hamper restoration projects in degraded peat areas where nature conservation tries to restore species-rich vegetation types with a high nature value.     

An improved method of partially digesting plant megabase DNA suitable for YAC cloning: application to the construction of a 5.5 genome equivalent YAC library of tomato

An improved method for preparing partially digested tomato DNA has been developed, that is suitable for YAC cloning. It involves (i) isolation of high molecular-weight DNA from agarose-embedded leaf protoplasts, (ii) controlled partial digestion in situ using Eco RI endonuclease in the presence of Eco RI methylase (M. Eco RI), and (iii) fractionation of the partial digest on a Clamped Homogeneous Electric Fields (CHEF) gel. Unlike methods commonly used for generating partial digests, the present method allows one to produce digests in which the bulk of restriction fragments are of the desired size. Use of these partial digests in constructing YAC libraries of the tomato lines Moneymaker- Cf4 and VFNT Cherry resulted in libraries (total 21 060 clones, 5.5 genome equivalents) in which 80% of the YACs have inserts between 200 and 600 kb. Both libraries have been screened with selected RFLP markers linked to the Cladosporium fulvum Cf4 locus on chromosome 1, using a three-dimensional PCR-based screening technique. To this end, the RFLP markers have been sequenced to allow for the synthesis of specific primers. Thus, for each marker tested several YAC clones have been isolated, including a family of clones that carry leucine-rich repeat sequences located around the Cf4/ Cf9 locus.     

Enhanced nitrogen loss may explain alternative stable states in dune slack succession

Ecological theory emphasizes competitive interactions between plant species when explaining primary succession in plants. Ecosystem processes, such as nutrient accumulation, are often regarded as independent, steering successional changes without being affected by the interacting plant species. We present experimental evidence that plant species in wet dune slack systems are able to affect ecosystem processes in their favor by reducing the rate of nitrogen accumulation by coupled nitrification–denitrification, promoting their competitive position. We compared denitrification rates of two early successional species having radial oxygen loss (ROL) with two non-ROL late successional species in a mesocosm experiment. The denitrification rates were significantly higher in mesocosms planted with early successional species Littorella uniflora (P<0.001) and Schoenus nigricans (P<0.05), relative to the rates found in the presence of non-ROL species, Carex nigra and Calamagrostis epigejos . We analyzed the consequences of enhanced nitrogen loss on the competition between an early and a late successional species by means of a simple theoretical model. Our analysis revealed that early successional species capable of ROL might retard successional changes and lock the ecosystem in an unproductive state for an extended period of time. We emphasize that understanding of ecosystem processes is crucial in successful conservation of high biodiversity vegetation in wet dune slacks.